Previously we have found that insulin in the concentration of 50 ng/ml when substituted for serum in Ham's F-12 medium, was sufficient to maintain the survival and differentiation of dissociated chick dorsal root ganglion neurons in vitro. Insulin, while necessary, was not sufficient to provide for the adequate survival of central neurons, chick spinal cord neurons in this serum-free medium. We are investigating the requirement of other hormones and/or growth factors for the survival of central neurons in culture. We are also currently investigating surface distribution and internalization of neuronal acetylcholine receptors using alpha-bungalotoxin as a specific ligand and chick ciliary ganglion neurons as target cells. Previously we have produced a series of monoclonal antibodies directed against neuronal surface antigens. Each of these antibodies will be characterized to detect the precise antigenic determination that the antibody is recognizing.